Search PubMed⌕ Search

Biomedical subjects

B Halliwell

Publications and source records attributed to B Halliwell.

At least 145 records · Page 8Linked to original sources
PubMed1997

Peroxynitrite-dependent aromatic hydroxylation and nitration of salicylate and phenylalanine. Is hydroxyl radical involved?

There is considerable dispute about whether the hydroxylating ability of peroxynitrite (ONOO-)- derived species involves hydroxyl radicals (OH.). This was investigated by using salicylate and phenylalanine, attack of OH. upon which leads to the formation of 2,3- and 2,5-dihydroxybenzoates, and o- m- and p- tyrosines respectively. On addition of ONOO- to salicylate, characteristic products of hydroxylation (and nitration) were observed in decreasing amounts with rise in pH, although added products of hydroxylation of salicylate were not recovered quantitatively at pH 8.5, suggesting further oxidation of these products and underestimating of hydroxylation at alkaline pH. Hydroxylation products decreased in the presence of several OH. scavengers, especially formate, to extents similar to those obtained when hydroxylation was achieved by a mixture of iron salts, H2O2 and ascorbate. However, OH. scavengers also inhibited formation of salicylate nitration products. Ortho, p- and m-tyrosines as well as nitration products were also observed when ONOO- was added to phenylalanine. The amount of these products again decreased at high pH and were decreased by addition of OH. scavengers. We conclude that although comparison with Fenton systems suggests OH. formation, simple homolytic fission of peroxynitrous acid (ONOOH) to OH. and NO2. would not explain why OH. scavengers inhibit formation of nitration products.

Chromatography, High Pressure Liquid↗
PubMed1996

Catalytic metal ions and the loss of reduced glutathione from University of Wisconsin preservation solution.

The University of Wisconsin solution is widely used for organ preservation. We show that this fluid is often contaminated by traces of iron catalytic for free radical reactions, as demonstrated by the bleomycin assay and the ability of the iron to stimulate lipid peroxidation. Reduced glutathione (GSH) added to University of Wisconsin solution during its commercial preparation had oxidized to the disulfide in all samples examined. Addition of GSH to UW solution showed its half-life to be about one day. The metal ion chelators desferrioxamine and phenanthroline did not substantially delay the loss of GSH, although desferrioxamine was able to suppress iron-mediated lipid peroxidation induced by the contaminant iron in the preservation solution. Additional iron was released from rat liver after a period of cold storage/rewarming.

Adenosine↗
PubMed1996

Do human atherosclerotic lesions contain nitrotyrosine?

Nitrotyrosine has been widely used as a marker of peroxynitrite formation in normal and diseased tissues. However, studies of normal human aortic intima and atherosclerotic lesions at various stages of development failed to reveal its presence except in a very few specimens of both normal and diseased tissues. The techniques used to detect nitrotyrosine-HPLC with photodiode array detection and Western blotting- were able to identify nitrotyrosine after chemical nitration. Hence peroxynitrite may not be as important in the initiation of atherosclerosis as has commonly been proposed.

Antibody Specificity↗
PubMed1996

Measurement of oxidized and methylated DNA bases by HPLC with electrochemical detection.

Oxidative DNA damage is thought to be an important contributor to cancer development and to be affected by dietary constituents, so its accurate measurement is important. DNA methylation is recognized as an important mechanism affecting gene expression. In the present paper we describe an HPLC-with-electrochemical-detection procedure that allows rapid and sensitive measurement of four oxidized (2,6-diamino-4-hydroxy-5-formamidopyrimidine, 5-hydroxyuracil, 8-hydroxyguanine, 8-hydroxyadenine) and three methylated (7-methylguanine, 1-methylguanine, O6-methylguanine) bases in acid hydrolysates of DNA. Guanine was also detected, but was clearly separated from the other bases.

Chromatography, High Pressure Liquid↗
PubMed1996

Formation of nitrating and chlorinating species by reaction of nitrite with hypochlorous acid. A novel mechanism for nitric oxide-mediated protein modification.

Detection of 3-nitrotyrosine has served as an in vivo marker for the production of the cytotoxic species peroxynitrite (ONOO-). We show here that reaction of nitrite (NO2-), the autoxidation product of nitric oxide (.NO), with hypochlorous acid (HOCl) forms reactive intermediate species that are also capable of nitrating phenolic substrates such as tyrosine and 4-hydroxyphenylacetic acid, with maximum yields obtained at physiological pH. Monitoring the reaction of NO2- with HOCl by continuous flow photodiode array spectrophotometry indicates the formation of a transient species with spectral characteristics similar to those of nitryl chloride (Cl-NO2). Reaction of synthetic Cl-NO2 with N-acetyl-L-tyrosine results in the formation of 3-chlorotyrosine and 3-nitrotyrosine in ratios that are similar to those obtained by the NO2-/HOCl reaction (4:1). Tyrosine residues in bovine serum albumin are also nitrated and chlorinated by NO2-/HOCl and synthetic Cl-NO2. The reaction of N-acetyl-L-tyrosine with NO2-/HOCl or authentic Cl-NO2 also produces dityrosine, suggesting that free radical intermediates are involved in the reaction mechanism. Our data indicate that while chlorination reactions of Cl-NO2 are mediated by direct electrophilic addition to the aromatic ring, a free radical mechanism appears to be operative in nitrations mediated by NO2-/HOCl or Cl-NO2, probably involving the combination of nitrogen dioxide (.NO2) and tyrosyl radical. We propose that NO2- reacts with HOCl by Cl+ transfer to form both cis- and trans-chlorine nitrite (Cl-ONO) and Cl-NO2 as intermediates that modify tyrosine by either direct reaction or after decomposition to reactive free and solvent-caged Cl. and .NO2 as reactive species. Formation of Cl-NO2 and/or Cl-ONO in vivo may represent previously unrecognized mediators of inflammation-mediated protein modification and tissue injury, and offers an additional mechanism of tyrosine nitration independent of ONOO-.

Hypochlorous Acid↗
PubMed1996

Oxidative DNA damage in human respiratory tract epithelial cells. Time course in relation to DNA strand breakage.

When human respiratory tract epithelial cells were exposed to 100 microM H2O2, there was rapid induction of DNA strand breakage and chemical modifications to all 4 DNA bases suggestive of attack by OH.. The major products were FAPy-adenine, FAPy-guanine, and 8-OH-guanine. Some of the base modifications were removed very quickly from the DNA (e.g., 8-OH-guanine), whereas others persisted for longer (e.g., thymine glycol), probably due to differential activity of different repair enzymes. By contrast, strand breaks continued to increase over the time course of the experiment, perhaps because strand breakage is also implicated in the repair process. One should therefore be cautious in using strand breakage as a sole measure of oxidative DNA damage, and when drawing conclusions about the pattern and biological significance of oxidative DNA damage in cells the relative persistence of different lesions must be considered.

Bronchi↗
PubMed1996

Protection against peroxynitrite-dependent tyrosine nitration and alpha 1-antiproteinase inactivation by oxidized and reduced lipoic acid.

Peroxynitrite, formed by combination of superoxide radical with nitric oxide, is a reactive tissue-damaging species apparently involved in the pathology of several human diseases. Peroxynitrite nitrates tyrosine residues and inactivates alpha 1-antiproteinase. We show that both lipoic acid and dihydrolipoic acid efficiently protect against damage by peroxynitrite. By contrast, other disulphides tested did not. The biological antioxidant effects of lipoate/dihydrolipoate may involve scavenging of reactive nitrogen species as well as reactive oxygen species.

Antioxidants↗
PubMed1996

An evaluation of the antioxidant and antiviral action of extracts of rosemary and Provençal herbs.

Extracts of herbs and spices are increasingly of interest in the food industry because they retard oxidative degradation of lipids. There is also increasing interest in the antiviral activity of plant products. A liquid, deodorized rosemary extract and an oily extract of a mixture of Provençal herbs were tested for antioxidant and antiviral action in vitro. The rosemary extract (Herbor 025) and the extract of Provençal herbs (Spice Cocktail) inhibited peroxidation of phospholipid liposomes with 50% inhibition concentration values of 0.0009% (v/v) and 0.0035% (v/v), respectively. Herbor 025 and the spice cocktail (at 0.2%, v/v) reacted with trichloromethylperoxyl radical with calculated rates of 2.7 x 10(4) s-1 and 1.5 x 10(3) s-1, respectively. The main active components in the herbal preparations, carnosol and carnosic acid, at 0.05% (v/v) react with rate constants of (1-3) x 10(6) M-1 sec-1 and 2.7 x 10(7) M-1 sec-1, respectively. Both extracts show good antioxidant activity in the Rancimat test, especially in lard. Herbor 025 and the spice cocktail inhibited human immunodeficiency virus (HIV) infection at very low concentrations which were also cytotoxic. However, purified carnosol exhibited definite anti-HIV activity at a concentration (8 microM) which was not cytotoxic. Both preparations promoted some DNA damage in the copper-phenanthroline and the bleomycin-iron systems. The two herbal preparations possess antioxidant properties that may make them useful in the food matrix.

Abietanes↗
PubMed1996

Evaluation of the antioxidant activity of melatonin in vitro.

Melatonin is being increasingly promoted as a treatment for "jet lag" and insomnia and has been suggested to act as an antioxidant in vivo. The antioxidant and potential pro-oxidant activities of melatonin were investigated in vitro. Melatonin was able to scavenge hypochlorous acid (HOCl) at a rate sufficient to protect catalase against inactivation by this molecule. Melatonin could also prevent the oxidation of 5-thio-2-nitrobenzoic acid by HOCl. Melatonin decreased the peroxidation of ox-brain phospholipids with a calculated IC50 of (210 +/- 2.3) microM. In contrast, serotonin which also scavenged HOCl, was much more effective in decreasing phospholipid peroxidation (IC50 15 +/- 5 microM). Both compounds reacted with trichloromethylperoxyl radical (CCl3O2) with rate constants of (2.7 +/- 0.2) x 10(8) and (1.2 +/- 0.1) x 10(8)M-1 s- respectively. Melatonin did not scavenge superoxide radical and weakly protected DNA against damage by the ferric bleomycin system. By contrast serotonin was weakly pro-oxidant in the ferric-bleomycin system and strongly pro-oxidant in the Fe(3+)-EDTA/H2O-deoxyribose system. Solubility restrictions precluded examination of melatonin in this system. Our data show that melatonin exerts only limited direct antioxidant activities.

Animals↗
PubMed1996

Blood radicals: reactive nitrogen species, reactive oxygen species, transition metal ions, and the vascular system.

Free radicals, such as superoxide, hydroxyl and nitric oxide, and other "reactive species", such as hydrogen peroxide, hypochlorous acid and peroxynitrite, are formed in vivo. Some of these molecules, e.g. superoxide and nitric oxide, can be physiologically useful, but they can also cause damage under certain circumstances. Excess production of reactive oxygen or nitrogen species (ROS, RNS), their production in inappropriate relative amounts (especially superoxide and NO) or deficiencies in antioxidant defences may result in pathological stress to cells and tissues. This oxidative stress can have multiple effects. It can induce defence systems, and render tissues more resistant to subsequent insult. If oxidative stress is excessive or if defence and repair responses are inadequate, cell injury can be caused by such mechanisms as oxidative damage to essential proteins, lipid peroxidation, DNA strand breakage and base modification, and rises in the concentration of intracellular "free" Ca(2+). Considerable evidence supports the view that oxidative damage involving both ROS and RNS is an important contributor to the development of atherosclerosis. Peroxynitrite (derived by reaction of superoxide with nitric oxide) and transition metal ions (perhaps released by injury to the vessel wall) may contribute to lipid peroxidation in atherosclerotic lesions.

Animals↗
PubMed1996

Hydroxyl radical generation by rheumatoid blood and knee joint synovial fluid.

OBJECTIVE: To demonstrate directly that highly reactive hydroxyl radicals (OH.) can be generated in patients with rheumatoid arthritis and contribute to joint damage, and to examine the ability of blood to cause OH. generation. METHODS: The sensitive and specific technique of hydroxylation of aromatic compounds (salicylate and phenylalanine) was used to measure OH.. Synovial fluid and blood from patients with active rheumatoid arthritis were aspirated and immediately added to tubes containing salicylate and phenylalanine as detectors of OH., or to tubes containing saline as a control. Levels of specific products of attack of OH. upon salicylate (2,3- and 2,5-dihydroxybenzoates) and phenylalanine (ortho- and meta-tyrosines) were measured by high performance liquid chromatography. RESULTS: Synovial fluid samples aspirated into saline never contained ortho- or meta-tyrosines or 2,3-dihydroxybenzoate. Of 53 patients examined, synovial fluid and blood from 36 caused formation of ortho- and meta-tyrosines when aspirated into solutions containing phenylalanine. Repeated sampling from three "positive" patients showed consistent evidence of these hydroxylation products. Similarly, of 22 patients examined, synovial fluid and blood from 18 caused formation of 2,3- and 2,5-dihydroxybenzoates when aspirated into salicylate solutions. Further evidence for the role of OH. was provided by inhibition of the hydroxylation by the specific OH. scavengers mannitol and sodium formate. CONCLUSIONS: Aspirated knee joint fluids and blood from rheumatoid arthritis patients can generate OH., consistent with current views on the importance of this radical as a cytotoxic agent in rheumatoid disease. The ability of body fluids to cause OH. formation is not correlated with simple laboratory indices of disease activity, but is reproducible on sequential sampling from the same patients. The mechanism and significance of the phenomenon in rheumatoid arthritis pathology remain to be established.

Arthritis, Rheumatoid↗
PubMed1996

Protection against peroxynitrite dependent tyrosine nitration and alpha 1-antiproteinase inactivation by some anti-inflammatory drugs and by the antibiotic tetracycline.

OBJECTIVE: To examine in vitro the ability of several drugs to protect against deleterious effects of peroxynitrite, a cytotoxic agent formed by reaction of nitric oxide with superoxide radical, that may be generated in the rheumatoid joint and could cause joint damage. METHODS: The ability of several drugs to protect against such possible toxic actions of peroxynitrite as inactivation of alpha 1-antiproteinase and nitration of tyrosine was evaluated. RESULTS: Most non-steroidal anti-inflammatory drugs were moderately (indomethacin, diclofenac, naproxen, tolmetin) or only weakly (sulindac, ibuprofen, aurothioglucose, flurbiprofen, sulphasalazine, salicylate, penicillamine disulphide) effective in preventing tyrosine nitration and alpha 1-antiproteinase inactivation by peroxynitrite, but 5-aminosalicylate and penicillamine were much more effective, as was the antibiotic tetracycline (but not ampicillin). Phenylbutazone and flufenamic acid protected effectively against tyrosine nitration, but could not be tested in the alpha 1-antiproteinase system. The analgesic paracetamol was highly protective in both assay systems. CONCLUSION: Many drugs used in the treatment of rheumatoid arthritis are unlikely to act by scavenging peroxynitrite. The feasibility of peroxynitrite scavenging as a mechanism of penicillamine, 5-aminosalicylate, and paracetamol action in vivo is discussed.

Acetaminophen↗
PubMed1996

Antioxidants in human health and disease.

Free radicals and antioxidants are widely discussed in the clinical and nutritional literature. Antioxidants are needed to prevent the formation and oppose the actions of reactive oxygen and nitrogen species, which are generated in vivo and cause damage to DNA, lipids, proteins, and other biomolecules. Endogenous antioxidant defenses (superoxide dismutases, H2O2-removing enzymes, metal binding proteins) are inadequate to prevent damage completely, so diet-derived antioxidants are important in maintaining health. Many dietary compounds have been suggested to be important antioxidants: The evidence for a key role of vitamins E and C is strong, but that for carotenoids and related plant pigments is weaker. Interest is also growing in the role of plant phenolics, especially flavonoids. Some antioxidants can exert prooxidant effects in vitro, but their physiological relevance is uncertain. Experimental approaches to the optimization of antioxidant nutrient intake are proposed.

Antioxidants↗
PubMed1996

Hydroxyl radical production during myocardial ischemia and reperfusion in cats.

We previously showed that generation of reactive oxygen species during myocardial ischemia and reperfusion stimulates cardiac sympathetic afferent nerve endings. We hypothesized that, in this feline model of brief ischemia and reperfusion, HO. is produced during ischemia and the rate and concentration of production of HO.during reperfusion is dependent on the duration of myocardial ischemia. Therefore, we evaluated the time dependency of production of HO. during reperfusion after 2, 5, and 10 min of reversible occlusion of the left anterior descending (LAD) coronary artery to induce ischemia in cats (n = 10). Blood samples collected from the coronary vein at 0.25, 1, 2, and 4 min after 2 min of ischemia revealed net cumulative rate of production of p-, m-, and o-tyrosine of 99 +/- 31, 10 +/- 5.1, and 0.8 +/- 0.2 nmol.min-1.g-1, respectively. After 5 min of ischemia, net cumulative rates of production of p-, m-, and o-tyrosine during reperfusion were 177 +/- 63, 74 +/- 26, and 1.6 +/- 0.8 nmol.min-1.g-1, respectively, whereas after 10 min of ischemia production rates were 153 +/- 42, 78 +/- 29, and 2.1 +/- 0.5 nmol.min-1.g-1, respectively. The highest rate of production of tyrosines was observed immediately after ischemia, perhaps indicating a washout of HO.-derived products that had accumulated in the myocardium during ischemia. To evaluate production of HO. during ischemia, deoxygenated saline (PO2 10 +/- 0.9 mmHg) containing phenylalanine was perfused into the ischemic coronary vascular bed through a cannula placed in the LAD (n = 16). Perfusate was collected from the coronary vein during the 10 min of ischemia. Net production of HO. during ischemia, measured by the production of p-, m-, and o-tyrosine, was 82 +/- 11, 6.6 +/- 0.4, and 1.7 +/- 0.3 nmol.min-1.g-1, respectively. Pretreatment with deferoxamine (10 mg/kg, n = 7) or dimethylthiourea (10 mg/kg, n = 6) decreased net production of HO. during ischemia and reperfusion. These results demonstrate that HO. is produced during brief ischemia and reperfusion, with the greatest amount being produced immediately after ischemia. Additionally, we show that the duration of brief ischemia determines the rate of production of HO. during reperfusion.

Animals↗
PubMed1996

Production of hydroxyl radicals in contracting skeletal muscle of cats.

Reactive oxygen species increase during exhaustive contraction of skeletal muscle, but characterization of the specific species involved and their rates of production during nonexhaustive muscle contraction have not been investigated. We hypothesized that the production rate of hydroxyl radical (.OH) increases in contracting muscle and that this rate is attenuated by pretreatment with deferoxamine (Def) or dimethylthiourea (DMTU). We measured the rate of production of .OH before, during, and after 5 min of intermittent static contraction of the triceps surae muscles in cats (n = 6) using the formation of p-, m-, and o-tyrosines by hydroxylation of phenylalanine. L-Phenylalanine (30 mg/kg i.v.) was administered to each animal 3 min before contraction. Blood samples were collected from the popliteal vein 1 min before contraction; 1, 3, and 4.5 min during contraction; and 1 min after contraction. During and after contraction, the cumulative production rates of p-, m-, and o-tyrosines were elevated by 42.84 +/- 5.41, 0.25 +/- 0.04, and 0.21 +/- 0.03 nmol.min-1.g-1, respectively, compared with noncontracting triceps surae muscles. Pretreatment with Def (10 mg/kg i.v.; n = 5) or DMTU (10 mg/kg i.v.; n = 4) decreased the cumulative rates of production of p-, m-, and o-tyrosines during and after contraction. Additionally, the rate of tyrosine production increased in proportion to the percentage of maximal tension developed by the triceps surae muscles. These results directly demonstrate that .OH is produced in vivo in the skeletal muscle of cats during intermittent static contraction and that production can occur before the onset of fatigue.

Animals↗